Citrus aurantium (bitter orange) extract: Safety assessment by acute and 14-day oral toxicity studies in rats and the Ames Test for mutagenicity.

The primary active constituent in bitter orange extract (BOE) is p-synephrine. This study assessed the safety of a BOE standardized to 50% p-synephrine following short-term exposure to rats and by the Ames Test. Following 5000 mg/kg of the extract orally to female rats all animals survived. Administration at 2000 mg/kg to female rats for four days yielded no signs of toxicity. Five male and five female rats were administered the BOE at 0, 250, 500, 1000 and 2000 mg/kg/day for 14 days. No significant effects were observed at any dose with respect to body weights, food intake, absolute and relative organ weights, hematology, clinical chemistry, and pathology. Two male rats died after 2000 mg/kg with gastrointestinal impaction at necropsy. During week two of 1000 mg/kg and 2000 mg/kg/day, rats exhibited transient signs of repetitive burrowing of heads in the bedding material (hypoactivity) for about 15 and 45 min, respectively. The no-observed-effect-level (NOEL) was 500 mg/kg/day. The mutagenic potential was assessed at and up to the limit dose of 5000 µg/plate in a Salmonella typhimurium reverse mutation (Ames) test, performed in duplicate as a pre-incubation assay in the presence and absence of metabolic activation (S9). The BOE did not induce an increase in the frequency of revertant colonies at any dose in the five tester strains, and was therefore non-mutagenic.

Probing the wound healing potential of biogenic silver nanoparticles.

OBJECTIVE: Silver nanoparticles (AgNPs) are known for their antimicrobial profile and wound healing activities. However, cytotoxicity and cosmetic abnormalities associated with silver pose a major challenge in their translation for therapeutic applications. Our objective was to develop biogenic AgNPs, using a single-step green synthesis, and to investigate their in vitro and in vivo behaviour as wound-healing agents. METHOD: AgNPs were prepared using the green synthesis approach with aqueous Bryonia laciniosa leaves extract. The AgNPs were then evaluated for physicochemical properties, stability, and antimicrobial and in vivo wound healing activities. RESULTS: Stable AgNPs with characteristic absorption at 408nm and 15±3nm particle size were generated via the active involvement of Bryonia laciniosa. No loss of stability was detected after 6 months at room temperature. Antibacterial activity was observed against both Gram-negative and Gram-positive bacteria with no cytotoxicity observed in vitro at a concentration of 200 µg/mL and effective cytokine modulation. In vivo wound healing experiments showed improved wound contracting ability in rats where, after 14 days, wound alleviation was 47.1±2.2% in the control groups, compared with 78.1±1.4% and 92.6±6.7% for a silver-based marketed cream and the AgNPs, respectively. CONCLUSION: The developed AgNPs proved to be superior wound healing agents owing to scarless healing with insignificant inflammation and toxicity. DECLARATION OF INTEREST: There were no external sources of funding for this study. The authors have no conflicts of interest to declare.

Enhancing activity and phospholipase A2 activity: two independent activities present in the enhancing factor molecule.

Enhancing factor (EF), a molecule that increases the binding of epidermal growth factor (EGF) to A431 cells, was first isolated in our laboratory from mouse intestines, and subsequently shown to be a secretory form of phospholipase A2 (PLA2) [Mulherkar, Rao, Wagle, Patki and Deo (1993) Biochem. Biophys. Res. Commun. 195, 1254-1263]. We had proposed earlier that EF increases the binding of EGF by first binding to its own cell-surface receptor [identified as a 100 kDa molecule; Mulherkar and Deo (1986) J. Cell. Physiol. 127, 183-188], and then by creating a binding site for EGF. However, due to its PLA2 activity, there was a possibility that EF, by its phospholipase activity could be unmasking cryptic EGF receptors on the cell surface, thereby increasing the number of binding sites for EGF. To test whether enhancing activity and phospholipase activity are independent of each other, a series of mutations were created using the full-length EF cDNA as a template, expressed in 293 cells and the mutant recombinant proteins checked for EF as well as PLA2 activities. Our studies have shown that one of the mutant EF proteins, lacking PLA2 activity, retains EF activity. This demonstrates unambiguously that EF and PLA2 activities are two independent activities in the same molecule. Mutation in the Ca2+-binding loop resulted in loss of EF activity, thereby demonstrating that EF activity is Ca2+-dependent. The N-terminal region of the EF molecule appears to be crucial for the enhancing activity.

Sequence analysis of full length cDNA for enhancing factor/phospholipase A2.

Enhancing factor (EF) protein was initially purified as a modulator of epidermal growth factor from small intestines of mouse. The cDNA sequence, obtained by RT-PCR, revealed that EF belonged to the non-pancreatic, phospholipase A2 (PLA2) family. This was the first report of the mouse PLA2. In the present paper we report the complete cDNA sequence of EF gene, in which the 5' sequence has been obtained by RAcE-PCR. The predicted amino acid sequence was computer analysed and the putative sites for enzyme action, calcium binding and heparin binding have been identified. The complete protein sequence of EF along with 16 aligned sequences were used to infer a phylogenetic tree. From this data the mouse EF was grouped with other membrane associated PLA2 with a bootstrap value of 98% indicating that it belonged to this class.

Mechanism-based screens in the discovery of chemotherapeutic antibacterials.

Numerous assays have been developed over the last 40 years for the detection of novel antibacterial metabolites. I have discussed many of the successful strategies and suggested some potential targets. Although the trend toward mechanism-based assays is relatively recent, it is clear that they have had a profound impact on screening in drug discovery. Often a mechanism-based assay requires construction of specific strains and verification of the antibacterial role of the selected target. Since the conception and development of a mechanism-based screen depends upon knowledge of the specific target and perhaps a compound that affects that target, it is implicit that mode of action studies on compounds discovered through random screening may subsequently lead to new mechanistic assays. While serendipity continues to play a crucial role in any screen, target-directed assays appear to be a worthwhile approach in antibacterial screening.

Nutritional enrichment of bakery products by supplementation with nonwheat flours.

Bakery products are important ready-to-eat processed foods. The nutritional quality of these products is low because of the inferior nutritional composition of wheat grain per se. This is further accentuated with the use of refined flours in their preparations. Nutritional composition of these products can be improved by using quality wheat for milling, increased extraction rates, air classification of flours to obtain protein-rich nonwheat flours and their products. The flours and protein products of legumes, oilseeds, other cereals, tubers, corn gluten and germ, and rice bran can be used effectively as vegetable protein sources for nutritional enrichment of the bakery products. In this article, recent literature concerning the nutritional composition of major bakery products, sources of vegetable proteins for product enrichment, and modifications in conventional processing methods to maintain the rheological and sensory properties of supplemented bakery products are reviewed critically.

Nutritional quality of some improved cultivars of cowpea.

The seeds of 13 improved cowpea cultivars were analysed for proximate composition, methionine, tryptophan and polyphenols. Considerable genetic variations were observed for the contents of protein, nonprotein nitrogen, limiting amino acids and polyphenols. The globulins contributed about 50% of the total seed proteins. A significant proportion of proteins of high protein cultivars was found to be insoluble. The proteins of most of the cultivars were found deficient in methionine and tryptophan. Methionine and tryptophan levels were negatively correlated with protein and positively correlated with carbohydrate contents. The cultivars with colored seed coats contained higher amounts of polyphenols. The difference in polyphenol content between various cultivars were due to seed coat polyphenols.

Effects of heat treatments and germination on trypsin inhibitor activity and polyphenols in jack bean (Canavalia ensiformis L. DC).

The application of dry heat to seeds and meal was not effective in inactivating the TI and reducing the polyphenol content. Soaking for 24 h followed by cooking for 20 min, was equally effective in destroying the TI activity. Germination of jack bean seeds for 40 h decreased the levels of TI and polyphenols by 31% and 35%, respectively.

Pigeonpea as an important food source.

Pigeonpea is an important source of proteins, carbohydrates, B-group vitamins, and certain minerals. India contributes over 90% of the pigeonpea production in the world where it is mostly consumed as dehusked splits or dhal. In African countries and Latin America, it is mainly consumed as canned peas. In this review, world production and distribution, genetic background, and biochemical and nutritional properties, storage and processing of pigeonpea are discussed. Future research needs to improve the utilization of pigeonpea as human food are also addressed.

Chemistry and technology of green gram (Vigna radiata [L.] Wilczek).

Green gram or mung bean (Vigna radiata [L.] Wilczek) is an important food legume grown under tropical and subtropical conditions. It is an excellent source of protein and is almost free from flatulence-causing factors. Because of this, green gram seeds are preferred for feeding babies and those convalescing. The seeds contain a higher proportion of lysine than any other legume seeds. The seeds are processed and consumed as cooked whole beans or splits (dhals), sprouts, immature seeds, and flour and are used in various recipes. The proposed work will incorporate available information on nutritional composition, processing, and utilization of green gram. The results reported in the literature on the above aspects of green gram will be analyzed critically, and future research needs will be defined to improve the utilization of green gram as human food.

Biochemistry and technology of chickpea (Cicer arietinum L.) seeds.

Chickpea is an important source of proteins, carbohydrates, B-group vitamins, and certain minerals, particularly to the populations of developing nations. India contributes over 75% of the chickpea production in the world where it is mostly consumed as dhal, whole seeds, and several types of traditional, fermented, deep fried, sweetened, and puffed products. In this review, the world production and distribution, genetic background, biochemical and nutritional quality, and developments in storage and processing technology of chickpea are discussed. Future research needs, to improve the utilization of chickpea as human food, are addressed.

Nutritional composition, processing, and utilization of horse gram and moth bean.

Horse gram and moth bean are the unexploited legumes of the tropics and subtropics grown mostly under dry-land agriculture. The chemical composition is comparable with commonly cultivated legumes. Like other legumes, these are deficient in methionine and tryptophan. Horse gram is an excellent source of iron and molybdenum. Comparatively, horse gram seeds have higher trypsin inhibitor and hemagglutinin activities and polyphenols than moth bean seeds. Dehusking, germination, cooking, and roasting have been shown to produce beneficial effects on nutritional quality of both the legumes. Both the legumes require prolonged cooking to obtain product of acceptable nature. A soak solution (1.5% NaHCO3 + 0.5% Na2CO3 + 0.75% citric acid) treatment has been shown to reduce cooking time and improve protein quality. Moth bean is mostly consumed as dhal or sprouts. The whole seeds of horse gram are generally utilized as cattle feed. However, it is consumed as a whole seed, sprouts, or whole meal by a large population in rural areas of southern India. Medical uses of these legumes have been discussed.

Winged bean in human nutrition.

Protein calorie malnutrition is prevalent in many developing countries of the tropics and subtropics. Improvement of protein supply to meet the demand of a growing population necessitates utilization of unconventional protein sources. Winged bean, a high protein crop, is one of the important underexploited legumes of the tropics. All the plant parts, viz., seeds, immature pods, leaves, flowers and tubers are edible. Mature seeds contain 29 to 37% proteins and 15 to 18% oil. It has fairly good amounts of phosphorus, iron, and vitamin B. Essential amino acid composition of winged bean is very similar to that of soybean. The fatty acid composition is very much comparable to groundnut. It contains relatively high amounts of behenic acid and parinaric acid. The trypsin inhibitor in winged bean has been shown to be heat resistant. Other toxic factors such as hemagglutinins and cyanide have also been reported. Winged bean seeds are hard to cook. Soaking of seeds in the Rockland's soak solution containing sodium bicarbonate, sodium carbonate, sodium chloride, and sodium pyrophosphate reduces cooking time significantly. The potential uses of this important crop in human nutrition and future research needs are discussed.

Chemical, biochemical, and biological significance of polyphenols in cereals and legumes.

Polyphenols in cereals and legumes have been receiving considerable attention largely because of their adverse influence on color, flavor, and nutritional quality. These compounds belong to the flavonoid and tannin groups and are mostly located in the seed coat or pericarp of the grains. The pearl millet flavonoids have been identified as C-glycosylflavones by the combined use of paper chromatography and UV spectroscopy. Although nontoxic, physiological and nutritional significance of these compounds occurring in high amounts in the pearl millet grain are still not clearly understood. In view of aesthetic quality, bleaching of the millet grains in acidic solution is recommended. A large proportion of current assays involves spectrophotometry of tannin or its chromogen and tannin-protein interaction. Sorghum and legume tannins have been characterized as condensed tannins. Several factors such as plant type, age of the plant or plant parts, stage of development, and environmental conditions govern the polyphenol contents in plants. Polyphenols are known to interact with proteins and form tannin-protein complexes leading to either inactivation of enzymes or making proteins insoluble. These are implicated in decreasing the activities of digestive enzymes, protein and amino acid availabilities, mineral uptake, vitamin metabolism, and depression of growth. Polyphenols are known to cause certain ultrastructural changes in the different parts of experimental animals. A correlation between dietary tannins and occurrence of esophageal cancer has been established. Bird resistance and seed germination in food crops have been correlated to high contents of polyphenols. The antinutritional activity of polyphenols can be reduced by removing polyphenols from the grains by chemical treatments or removing pericarp and testa by pearling. Treatment of alkaline reagents and ammonia can remove 90% of the polyphenols. Supplementation of polyphenols-rich diet with protein can alleviate the growth-depressing effect of polyphenols.